Description

Mechanisms of CFTR activation by S-nitrosoglutathione (normal and CF)

Cystic fibrosis (CF) is a multisystem disease associated with mutations in the gene
encoding the CF transmembrane conductance regulatory ( CFTR ) protein [1]. CFTR has several functions but is typically regarded as an apical
membrane Cl - channel in epithelial cells. Its post-translational processing
involves complex and incompletely defined series of interactions with variety of
chaperones and co-chaperones that assist in proper folding, of CFTR, as well as
its glycosylation, and assess the folded protein it for possible defects. The most common
mutation associated with CF, deltaF508, results in a single amino acid deletion [2], [3]. The majority of wild-type (wt) CFTR, and virtually
all deltaF508 CFTR, is degraded before reaching the cell surface [4], [5]. Certain agents and conditions increase expression, maturation, and
function of deltaF508 CFTR[6].

S-Nitrosoglutathione is an endogenous bronchodilator and signaling
molecule [7] that enhances expression, maturation, and function of both wt
and deltaF508 CFTR in epithelial cells [8], [9],
[10]. S-Nitrosoglutathione is present endogenously on the apical side
of airway epithelium. It increases ciliary beat frequency, thereby improving mucociliary
clearance [11].

Nitric oxide synthases (NOSs) are involved in conversion of L-arginine into
nitric oxide ( NO ). NO, in turn, is involved in production of
S-nitrosothiols, including S-Nitrosoglutathione. NO may react directly
with thiyl radicals or with thiols to form S-nitrosothiols or S-nitrosothiol radicals,
respectively [12], [13], [14].

In human, there are three isoforms of NOS: neuronal NOS ( nNOS ), endothelial
NOS ( eNOS ) and inducible NOS ( iNOS ). nNOS and eNOS are
constitutively expressed and produce small amounts of NO, whereas iNOS
expression is mainly induced by inflammatory stimuli. Induced iNOS synthesizes relatively
large quantities of NO. All three isoforms are expressed in human airways [13].

In the case of CF, airway epithelial cells are more susceptible to bacterial and viral
infection due to impairment of the host NO defense pathway. Polymorphisms of constitutive
NOS ( nNOS and eNOS ) and reduced iNOS expression contributes to
decreased NO production along with bacterial consumption [15], [16], [17], [13], [18].

S-nitrosylation can functionally regulate the general activities of Heat shock protein
90kDa alpha ( HSP90 alpha ) and provide a feedback mechanism for limiting
eNOS activation. S-Nitrosoglutathione covalently modifies a susceptible
cysteine residue in the HSP90 alpha domain that interacts with eNOS. On
the one hand, S-nitrosylation abolishes the positive regulation on eNOS activity
mediated by native chaperone HSP90 alpha[19]. On the other hand,
direct S-nitrosylation can increase the activity of each of the major forms of nitric
oxide synthases ( nNOS, eNOS and iNOS ) [20].

Ceruloplasmin also may catalyze the synthesis of S-Nitrosoglutathione[21], [22], [14].

Gamma GT can be involved in CFTR activation.
S-Nitrosocysteinylglycine, the product of S-Nitrosoglutathione cleavage by
Gamma GT, can increase DeltaF508 CFTR maturation [29].

SOD1, TXNRD1, TXNRD2 and TXNRD3 catabolize
S-Nitrosoglutathione to form free NO radicals [23], [28], [22], [11]. Free NO can spontaneously react
with Superoxide anion radical ( O(2)(-) ) to produce Peroxynitrite (
ONOO(-) ) [30]. The presence of SOD1, that catalyzes the
dismutation of O(2)(-), can outcompete the peroxynitrite reaction. Cells may contain
sufficient SOD1 to prevent inactivation of NO by O(2)(-)[12].

S-Nitrosoglutathione also increases expression of DnaJ homolog,
subfamily C, member 5 ( Csp ) to enhance the association between Csp and
CFTR in the ER and Golgi. S-Nitrosoglutathione does not S-nitrosylate
Csp. S-Nitrosoglutathione actually increases Csp expression
(primarily post-transcriptionally) leading to increase in CFTR folding and
maturation [29].

In the absence of S-Nitrosoglutathione, Csp initiates activation of
HSC70 ATPase activity, which leads to CFTR degradation [34],
[35], [36]., This degradation is inhibited in the presence of
S-Nitrosoglutathione, allowing increased Csp to continue stabilization of
CFTR. HSC70 has a single critical cysteine residue in its ATP binding
domain. S-nitrosylation of this cysteine allows Csp to augment CFTR folding
without leading to CFTR degradation [29].